A core factor of PROTAC molecules is that protein degradants can be designed rationally by selecting ligase molecules and linkers according to the target. The module that binds to E3 ligase can be replaced for different targets to recruit E3 ligase with the best therapeutic effect.
Although PROTAC molecular designs based on recruiting CRBN and VHL ligases have become increasingly mature, their preclinical and clinical development stillPROTAC faces multiple challenges. Especially in the field of oncology, drug resistance can emerge rapidly, and tumors may evade the role of protein degraders dependent on non-essential ligases (such as CRBN and VHL) by producing genetic mutations. Preclinical studies found that some tumors can evade from protein degradants by introducing mutations or reducing expression in the components of the ubiquitin ligase system. These challenges urge the discovery of new E3 ligases.
E3 ligases that can be used in clinical development can be elaborated from the following aspects.
- Ubiquitous ligase
Like CRBN or VHL, these ligases can match any target protein and can be used to treat different diseases. At present, only a small group of ligases of this type are used in PROTAC molecules, which can be queried online in the database PROTAC-DB.
Interestingly, recruiting VHL or CRBN for the same target protein may lead to different degradation efficiency. For example, in the development of proteolytic agents for KRAS G12C, previous studies found that KRAS G12C can be targeted by the PROTAC system, but the degradation effect will be affected by whether the recruited E3 ligase is VHL or CRBN.
Several factors may affect the degradation characteristics of ligases, including the complementarity between the conformation of ligases and target proteins, the ability of ligases and target proteins to directly form ternary structures for degradation, the intracellular localization of different ligases and target proteins, and the expression of cell type-specific ligases and targets.
- Tissue and cell specific E3 ligase
The tissue and cell specificity of ligase, the degree of enrichment in tumors, and whether it is essential for tumors may present opportunities for the development of domain-specific proteolytic therapy. Both academia and pharmaceutical industries are interested in discovering E3 ligases with unique expression characteristics, so as to carry out precision targeted protein degradation (precision TPD).
Previous studies found that some E3 ligases such as RNF182 and TRIM9 are specifically expressed in the central nervous system (CNS), which may play an important role in targeting neuronal diseases. Specific targeted degradation of CNS can avoid systemic miss and side effects.
Interestingly, some ligases have particularly low levels of expression in specific tissues and cells, which may also provide opportunities for protein degradants. An excellent example is the low expression of VHL in platelets, so the PROTAC molecule DT2216 recruiting VHL can avoid harming platelets when it degrades BCL-XL, thereby reducing platelet toxicity and enhancing its therapeutic index. At present, it is in phase 1 clinical development.
- E3 ligase enriched in diseased tissues or cells
Another frontier of precise TPD is the PROTAC molecules that specifically target tumor cells. One strategy is to target tumor-specific or tumor-enriched E3 ligases. In general, E3 ligases enriched in tumors overlap with tumor-dependent E3 ligases. This correlation can be used to identify E3 ligases for many cancer cell types, with an advantage that tumor cells are difficult to produce resistance to PROTAC molecules through ligase mutations.
However, whether these ligases can really improve the therapeutic index of PROTAC molecules still needs to be validated. One problem is that most of the highly enriched or tumor-dependent E3 ligases in tumors are related to the cell cycle, and PROTAC molecules based on them may produce toxicity similar to that of classical chemotherapy drugs, but this still needs to be verified by experiments.
Exploring tumor-specific E3 ligase for protein degradation is only one of the strategies to obtain tumor-specific PROTAC molecules. Another strategy is to couple degradable molecules to tumor-specific antibodies. Studies from Genentech and its partners showed the tumor targeting specificity of antibody-coupled PROTAC molecules. However, antibody-coupled PROTAC loses the oral characteristics of small PROTAC.